Unbiased screen of human transcriptome reveals an unexpected role of 3′UTRs in translation initiation

Abstract

Although most eukaryotic mRNAs require a 5ʹ-cap for translation initiation, some can also be translated through a poorly studied cap-independent pathway. Here we developed a circRNA-based system and unbiasedly identified more than 10,000 sequences in the human transcriptome that contain Cap-independent Translation Initiators (CiTIs). Surprisingly, most of the identified CiTIs are located in 3ʹUTRs, which mainly promote translation initiation in mRNAs bearing highly structured 5ʹUTR. Mechanistically, CiTI recruits several translation initiation factors including eIF3 and DHX29, which in turn unwind 5ʹUTR structures and facilitate ribosome scanning. Functionally, we showed that the translation of HIF1A mRNA, an endogenous DHX29 target, is antagonistically regulated by its 5ʹUTR structure and a new 3ʹ-CiTI in response to hypoxia. Therefore, deletion of 3ʹ-CiTI suppresses cell growth in hypoxia and tumor progression in vivo. Collectively, our study uncovers a new regulatory mode for translation where the 3ʹUTR actively participate in the translation initiation.

Disruption of HIF1Α translational control attenuates the HIF-dependent hypoxic response and solid tumour formationin vivo

“…Our work has shown that the 5’UTR of HIF1α mRNA is necessary for the HIF-dependent hypoxic response by regulating HIF1α mRNA association with polysomes and regulatory proteins such as YB-1. In addition other work has shown the 3’ UTR of HIF1α also plays a positive role in regulating HIF1α mRNA translation rates by recruiting several translation initiations factors, including eiF3 and DHX29, which unwind the 5’UTR during translation initiation [22]. This work indicates that the 5’ and 3’ HIF1α UTRs act in concert to control HIF1α protein translation, indicating that examining the roles of UTRs in the context of an artificial reporter construct may miss the full picture of regulation [23].…”

“…Several studies using artificial reporter constructs have indicated that both the 5’ and 3’ UTRs of HIF1α can modulate translation rates through acting as internal ribosome entry sites (IRES), sites to recruit RNA binding proteins, or act as targets for miRNAs [13, 15]. However, the primary role for the endogenous HIF1α 5’UTR remains unclear, with conflicting studies using reporter constructs suggesting it represses translation through its complicated secondary structure [22], or activates translation by binding to specialised RNA-binding proteins to facilitate ribosome recruitment [15, 21]. As more recent work using genome editing approaches has indicated that reporter constructs investigating the role of UTRs do not always phenocopy the role of the endogenous UTRs in cells [23], we decided to modify the endogenous HIF1α 5’UTR in cells to investigate its function.…”